The proximal tubule reabsorbs low molecular weight protein (<70 kDa) that passes the glomerular filtration barrier by a mechanism that is largely dependent on the scavenger receptor megalin. In the healthy adult this amounts to as much as 0.6-5.0 g of protein per day. However in renal disease, where the glomerular barrier becomes damaged, protein delivery to the proximal tubule increases dramatically and reaches levels that are eventually cytotoxic. Under these conditions the endocytic capacity of the proximal tubule becomes saturated and results in progressive renal damage (nephrotic syndrome). In addition to proteinuria and sodium retention, the nephrotic syndrome is characterized by up-regulation of several vasoactive and inflammatory genes. Although the pathologic course of the nephrotic syndrome has been well characterized, the molecular mechanisms that link protein absorption with the disease as well as with gene regulation in the proximal tubule are not understood. Recent studies in our laboratory have shown that megalin is proteolytically processed by a series of enzymes that are similar, or identical to, those shown to act on other receptors and that are involved in important signaling pathways in other cell types. These processing events have been shown to link protein absorption with cellular signaling pathways. Our data suggests that megalin is involved in a similar, here-to-fore unknown, pathway in the proximal tubule. In order to further investigate megalin's role in renal protein absorption especially as it relates to signaling events we will utilize newly developed antibodies to megalin and carry out experiments using normal rat kidneys and a proximal tubule cell line called OKP. We will characterize in detail each of the megalin-specific proteolytic activities. We will describe the downstream signaling events mediated by megalin processing and identify genes that are regulated. We will also examine the role of megalin-associated. proteins including, NHE3, Dab2, ARM and myosin VI. Finally, we will examine the effect of chronic proteinuria on megalin-mediated absorption and signaling by studying these processes in the puromycin aminonucleoside-induced nephrotic rat kidney.